Abstract

We have previously reported that functionally active μ-opioid receptors (MOR) are constitutively expressed at relatively low levels by developing T cells in the thymus. However, very little is known about the regulation of MOR expression by immature T cells. In this report, we first attempted to determine the effect of T cell receptor-induced T cell activation on the expression of MOR. We activated T cells with either the combination of anti-CD3 and CD28, or with superantigen, and observed a substantial increase in MOR transcript expression. We also chose to examine the effect of cytokine-mediated T cell activation on the expression of this opioid receptor. We selected certain cytokines that play a role in T cell development and are known to be present at functional levels in the thymus gland. Our results show that interferon γ (IFNγ), IL-1β, and IL-2, and in particular transforming growth factor-β (TGFβ), all induced significant increases in MOR transcript expression. On the other hand, both TNFα and IL-7 exhibited much weaker effects on MOR expression. These results show that MOR expression by developing T cells is strongly regulated by several cytokines involved in T cell development in the thymus gland.

Analysis of the effects of cytokines on MOR expression in thymocytes. Panel A. Cells were treated with IL-1β at 0, 10, 50, 100, 200, or 400 U/ml (Lanes 1–6). Panel B. Cells were treated with TGFβ at 0, 0.02, 0.25, 2.5, 5, or 10 U/ml (Lanes 1–6). Panel C. Cells were treated with IFNγ at 0, 1, 10, 50, 100, 200 U/ml (Lanes 1–6). Following treatment with the designated doses of cytokine for 24 hr, the expression of MOR and G3PDH were determined. Results are representative of at least 4 experiments. Panel D. Cells were treated with the designated doses of cytokine for 24 hr, and the normalized MOR expression was determined. Results are presented as the mean (± SE) of at least 4 experiments. All data points are statistically significant (compared with 0 U/ml) except for IL-1 at 10 U/ml. Panel E. Increased inhibition of forskolin-stimulated cAMP accumulation by DAMGO administration in TGFβ-treated thymocytes. Thymocytes were cultured with or without TGFβ (5U/ml) for 24 hr and then subjected to analysis of the inhibition of forskolin-induced cAMP levels with graded doses of DAMGO. The level of cAMP in cell lysates was determined by ELISA as described in the Materials and Methods section. Results are presented as the mean (± SE) of 3 experiments. Data points for DAMGO doses above 1 nM are statistically significant (P < 0.05) comparing the TGFβ-treated and non-treated groups.